Novel combination of glucocorticoids and pde-4 inhibitors for treating respiratory diseases, allegic diseases, asthma and copd

ABSTRACT

The invention relates to a novel combination of a glucocorticoid, especially loteprednol, and at least one phospho-diesterase-4 inhibitor (PDE-4-inhibitor), especially hydroxyindole-derivative N-(3,5-dichloropyridine-4-yl)-2-[1-(4-fluorbenzyl)-5-hydroxyindole-3-yl]-2-oxoacetamide, for a simultaneous, sequential or separate administration in the treatment of respiratory diseases, allergic diseases, asthma and chronic obstructive pulmonary diseases (COPD).

The present invention relates to a novel combination of aglucocorticoid, especially loteprednol, and at least onephosphodiesterase-4 inhibitor (PDE-4 inhibitor), especially thehydroxyindole derivativeN-(3,5-di-chloropyridin-4-yl)-2-[1-(4-fluorobenzyl)-5-hydroxyindol-3-yl]-2-oxoacetamide,for a simultaneous, sequential or separate administration in thetreatment of respiratory diseases, allergic diseases, asthma and chronicobstructive pulmonary diseases (COPD).

Allergic diseases and chronic obstructive pulmonary diseases (COPD) arebased on inflammatory processes characterized by an increased number ofinflammatory cells and increased release or secretion of inflammationmediators. Studies over the last 20 years have revealed thatinflammation of the respiratory tract is of central importance for therespiratory dysfunction in asthma and COPD. Comparable changes have beenobserved in allergic inflammations of the nose and of the eyes.Normally, the mucosa is infiltrated by a large number of cells,including mast cells, eosinophils and lymphocytes. These cells release anumber of mediators, including in particular interleukin-4 (IL-4),GM-CSF (granulocyte/macrophage colony-stimulating factor) and the tumornecrosis factor α (TNF-α), which eventually bring about theinflammations and the symptoms of allergic diseases and of COPD.

At the present time, a similar anti-inflammatory therapeutic approach isfollowed for all allergic diseases. The pathology of these diseases hasrevealed that the inflammatory process in the mucosa of patientsprimarily determines the symptom activity. Of the anti-inflammatorycompounds currently available for the treatment of asthma, rhinitis orconjunctivitis, glucocorticoids are the most effective. Activeingredients which can be administered topically by inhalational,intranasal or intraocular administration are preferably employed. On thebasis of the successful use of inhalable glucocorticoids in thetreatment and prevention of respiratory inflammations and permanent lungdamage in asthma patients, this therapeutic approach has also beenapplied to COPD patients although there are no data which mightunambiguously prove a long-term efficacy of these active ingredients inCOPD patients (Whittaker A J, Spiro SG; Curr Opin Pulm Med 2000;6:104-9).

One of the most important anti-inflammatory properties ofglucocorticoids arises from inhibition of cytokine release. It is knownthat several cytokines such as IL-4, IL-5, GM-CSF and TNF-α are involvedin respiratory inflammation. The efficacy of glucocorticoids can in partbe explained by the inhibitory effect on cytokine synthesis and cytokinerelease (Marx et al.; Pulm Pharmacol Ther 2002; 15:7-15).

One disadvantage of glucocorticoids arises from their possible systemicside effects such as, for example, growth retardation or elseosteoporosis. Sensible measures for reducing the risk of side effects ontopical administration of glucocorticoids include the use of the minimumeffective dose or restriction of the systemic availability of the activeingredient. A novel route is opened up by the use of so-called softsteroids. In contrast to other glucocorticoids, most of which undergodegradation to pharmacodynamically inactive metabolites only in theliver, the soft steroids undergo partial metabolic inactivation even atthe site of their administration (intranasal, ocular or intrapulmonary).Following this partial local metabolism, only very little, or no,pharmacodynamically active substance reaches the systemic bloodcirculation, so that the steroid-specific side effects are not to beexpected in practice. The most prominent example of this novel class ofactive ingredients is loteprednol, which is already approved for thetherapy of allergic conjunctivitis and uveitis.

A further class of potential therapeutics for allergic diseases and COPDcomprises the phosphodiesterase-4 inhibitors. Phosphodiesterase enzymesare responsible for the inactivation of cyclic adenosine monophosphate(cAMP) and cyclic guanosine monophosphate (cGMP). Inhibition ofphosphodiesterase-4 leads to an increase in cAMP in the cells, in turnleading to downregulation of the function of virtually allproinflammatory cells or immune cells. It is of interest thatinflammatory cells involved in the pathogenesis of diseases such asasthma, conjunctivitis, rhinitis or chronic obstructive pulmonarydisease preferentially express the phosphodiesterase-4 enzymes.

In recent years there have been advances in the development ofphosphodiesterase-4 inhibitors which can be employed for the therapy ofallergic diseases, asthma or COPD. It has been possible to show the invitro inhibitory activity on cytokine release and the therapeuticefficacy in asthma models for example for the active ingredientsroflumilast, cilomilast or else piclamilast (Torphy et al.; PulmPharmacol Ther 1999; 12:131-5; Poppe et al.; Allergy 2000; 55(Suppl63):270; Giembycz M A; Expert Opin Investig Drugs 2001; 10:1361-79;Ezeamuzie C I; Eur J Pharmacol 2001; 417:11-8). There is particularinterest in a novel class of substituted hydroxyindoles which aredescribed in DE 19 818 964, DE 19 917 504 and U.S. Pat. No. 6,251,923,and also novel 7-azaindoles which are disclosed in DE 10 053 275 andPCT/EP 01/12376.

It has now surprisingly been found that the novel combination of aglucocorticoid with at least one phosphodiesterase-4 inhibitor isadvantageous in the treatment of respiratory diseases, allergicdiseases, asthma and/or chronic obstructive pulmonary diseases. Add-ontherapy of a phosphodiesterase-4 inhibitor, especially the hydroxyindolederivativeN-(3,5-dichloropyridin-4-yl)-2-[1-(4-fluorobenzyl)-5-hydroxyindol-3-yl]-2-oxoacetamide,which can be administered orally, intranasally or by inhalation, withtopical glucocorticoids, especially loteprednol, is distinguished byimproved therapeutic efficacy as well as by the occurrence of few sideeffects.

The invention serves to improve the therapy of respiratory diseases,allergic diseases, asthma and chronic obstructive pulmonary diseases, aswell as the prophylaxis thereof. It is possible with aphosphodiesterase-4 inhibitor present in the combination and with aglucocorticoid successfully to control the inflammations which underliethe pathological states. Moreover, add-on therapy withphosphodiesterase-4 inhibitor leads to a smaller use of glucocorticoids,thus reducing the risk of side effects.

The present invention therefore relates to a composition which comprisesa glucocorticoid and at least one phosphodiesterase-4 inhibitor in fixedor free combination, and to the use thereof for producing a medicament.The invention also relates to a medicament for the treatment ofrespiratory diseases, allergic diseases, asthma and/or chronicobstructive pulmonary diseases, which comprises as active ingredient aglucocorticoid and at least one phosphodiesterase-4 inhibitor in fixedor free combination, and to a process for the production thereof.

It is possible to employ all glucocorticoids for the purposes of thepresent invention. So-called soft steroids are preferably used. Theexamples which may be cited of glucocorticoids which can be employedaccording to the invention are beclomethasone(9-chloro-11β,17,21-trihydroxy-16β-methyl-1,4-pregnadiene-3,20-dione),especially beclomethasone dipropionate, budesonide(16α,17-butylidenedioxy-11β,21-dihydroxy-1,4-pregnadiene-3,20-dione),ciclesonide (see, for example, WO 98/52542 and literature citedtherein), fluticasone (S-(fluoromethyl) 6α,9-difluoro-11β-carbothioate),especially fluticasone propionate, mometasone(9,21-dichloro-11β,17-dihydroxy-16α-methyl-1,4-pregnadiene-3,20-dione),in particular mometasone furoate, and loteprednol, especiallyloteprednol etabonate (chloromethyl17α-[(ethoxycarbonyl)oxy]-11β-hydroxy-3-oxoandrosta-1,4-diene-17β-carboxylate).

In a preferred embodiment of the invention, loteprednol and itspharmaceutically acceptable esters, especially loteprednol etabonate, isused as soft steroid. The preparation of loteprednol and loteprednoletabonate is described for example in the German patent DE 3 126 732,the corresponding U.S. Pat. No. 4,996,335 and the corresponding Japanesepatent JP-89011037.

Further soft steroids suitable according to the invention are describedfor example in the German patent DE 3 786 174, the corresponding patentEP 0 334 853 and the corresponding U.S. Pat. No. 4,710,495.

It is possible for the purposes of the present invention to employ allphosphodiesterase-4 inhibitors. These include, in particular but notrestrictively, the class of substituted hydroxyindole derivatives whichare described in DE 19 818 964, DE 19 917 504 and U.S. Pat. No.6,251,923, and also novel 7-azaindole derivatives which are disclosed inDE 10 053 275 and PCT/EP 01/12376. Examples of phosphodiesterase-4inhibitors which can be used according to the invention are rolipram((R)-4-[3-(cyclopentyloxy)-4-methoxyphenyl]-2-pyrrolidinone),roflumilast (Byk-Gulden), piclamilast (Rhone-Poulenc Rorer), cilomilast(GlaxoSmithKline) and the hydroxyindole derivativeN-(3,5-dichloropyridin-4-yl)-2-[1-(4-fluorobenzyl)-5-hydroxyindol-3-yl]-2-oxoacetamide.Particular preference is given to the substituted hydroxyindolederivativeN-(3,5-dichloropyridin-4-yl)-2-[1-(4-fluoro-benzyl)-5-hydroxyindol-3-yl]-2-oxoacetamide(“DFHO” hereinafter), which is described for example in DE 19 818 964.The phosphodiesterase-4 inhibitors can also be employed aspharmaceutically acceptable salts as are known to the skilled worker.

The inventive combination of a glucocorticoid, in particular of a softsteroid, with at least one phosphodiesterase-4 inhibitor can beadministered both prophylactically and after appearance of symptoms.They can also be used to retard or prevent progression of the diseases.

In a preferred embodiment, a combination of the active ingredientsloteprednol etabonate andN-(3,5-dichloropyridin-4-yl)-2-[1-(4-fluorobenzyl)-5-hydroxy-indol-3-yl]-2-oxoacetamide(DFHO) is used.

The following description of experiments serves to explain the inventiveteaching in detail without restricting it.

Inhibition of GM-CSF Release from LPS-Stimulated Monocytes

EDTAized human whole blood was mixed with Hanks' buffer in the ratio1:1. Histopaque 1077 solution (15 ml) was cautiously overlaid with max.40 ml of the blood: Hanks' mixture and centrifuged (2000 rpm) at roomtemperature for 30 min. The band enriched with leukocytes was aspiratedoff, washed twice with Hanks' buffer and transferred into RPMI 1640medium with Glutamax I (Gibco BRL, Eggenstein). The monocytes wereremoved through their adherence to the cell culture bottle over a periodof two hours. The cells were then thoroughly washed with medium in orderto remove non-adherent cells. The resulting monocytes were cultured inRPMI 1640 medium with 10% heat-inactivated fetal calf's serum (FCS) and100 U/ml penicillin and 100 μg/ml streptomycin in a CO₂ incubator (5%CO₂, 96% relative humidity, 37° C.).

Primary monocytes were seeded in 24-well plates at 5×10⁵ cells/well. Thecells were preincubated with the stated test substances for 30 minutes.LPS was then added, and incubation was continued for a period of 24 h.The supernatants were aspirated off and investigated by ELISA.

The amount of secreted human GM-CSF in the cell culture supernatants wasdetermined by using an OptEIA™ human GM-CSF ELISA test (Pharmingen, SanDiego). It was carried out in microtiter plates. Anti-human monoclonalantibodies were coupled as antibodies to the plate at 4° C. overnight.This coating and three washes were followed by saturation of nonspecificbindings by means of assay diluent solution™ (PBS with 10% FCS, pH 7.0)(Pharmingen, San Diego) at RT for 1 h. This was followed by incubationwith the samples and the standard (recombinant human GM-CSF) at 4° C.overnight. The samples were prepared undiluted or in a dilution of 1:50,of, the standard dilutions according to the protocol starting from astock solution with 500 pg/ml human GM-CSF. Bound human GM-CSF wasdetected with the aid of biotinylated monoclonal anti-human GM-CSFantibodies and an avidin-horseradish peroxidase reagent at RT for 1 h.All the steps were followed by washing 5 or 7 times with PBS/0.05% Tween20. The enzyme activity was determined using substrate solution™(tetramethylbenzidine (TMB) and hydrogen peroxide, Pharmingen, SanDiego) as substrate at RT for 30 min. The enzyme-substrate reaction wasstopped with 1M phosphoric acid, and the extinction at 450 nm wasmeasured.

Results

Firstly, dose-activity plots were established separately forN-(3,5-dichloropyridin-4-yl)-2-[1-(4-fluorobenzyl)-5-hydroxyindol-3-yl]-2-oxoacetamide(DFHO) and loteprednol. From these, the IC₅₀ for GM-CSF release fromhuman monocytes was calculated respectively as 3.2 μM for DFHO and 53.7nM for loteprednol. In further experiments, IC₅₀ values for DFHO andloteprednol were established in the presence of sub-IC₅₀ concentrationsof the respective other substance. In these cases, addition of 5 nM DFHOlowered the IC₅₀ for loteprednol from 53.7 nM to 13.4 nM. Conversely,addition of 10 nM loteprednol lowered the IC₅₀ for DFHO from 3.2 μM to0.06 μM. The IC₅₀ values found for loteprednol for release of TNF and ofGM-CSF from LPS-stimulated monocytes correspond to the IC₅₀ valuesindicated in the literature for other cell systems. This means that thecell system used is valid and suitable, and the investigations which arenecessary for the aim of the project with this system come to a reliableconclusion. The IC₅₀ values for DFHO correspond to those valuesindicated in the patent literature.

When 5 nM DFHO was given, the reduction in the IC₅₀ for loteprednol forTNF release was 65% and for GM-CSF release was 75%. The concentration of5 nM DFHO is far below the IC₅₀ for this substance, which isrespectively 5.7 μM and 3.2 μM, so that no effect is to be observed when5 nM DFHO is given on its own.

Conversely, the reduction in the IC₅₀ for DFHO for TNF release was 99%and for GM-CSF release was 98% when 10 nM loteprednol was givensimultaneously. The concentration of 10 nM loteprednol is far below theIC₅₀ of this substance, which is 85.5 nM and 53.7 nM respectively, sothat no effect is to be observed when 10 nM loteprednol is given on itsown.

A surprising observation which could not have been predicted by theskilled worker is that there is here a superadditive effect broughtabout by the simultaneous administration of loteprednol and DFHO on theinhibition of TNF and GM-CSF release.

The dosage forms mentioned below are particularly suitable foradministration of the inventive combination of active ingredients.

Thus, the active ingredients present in the combination can for examplebe administered separately as two oral formulations, or one activeingredient is in the form of an oral formulation and the other is intopical form (intranasal, inhalational).

In one embodiment of the invention, the phosphodiesterase-4 inhibitorcan be administered orally. Customary pharmaceutical formulations areused in this case, such as tablets, syrup, capsules, preparations withslowed release (sustained release formulation), pastilles oreffervescent granules.

Solid pharmaceutical forms such as tablets may comprise inertingredients and carriers such as, for example, calcium carbonate,calcium phosphate, sodium phosphate, lactose, starch, mannitol,alginates, gelatin, guar gum, magnesium stearate or aluminum stearate,methyl-cellulose, talc, colloidal silicas, silicone oil, high molecularweight fatty acids (such as stearic acid), agar-agar or vegetable oranimal fats and oils, solid high molecular weight polymers (such aspolyethylene glycol); preparations suitable for oral administrationsmay, where appropriate, comprise additional flavorings or sweeteners.The compositions in capsule form can be produced by generally customaryprocesses, for example by using the aforementioned carriers in a hardgelatin capsule shell. For compositions in the form of soft gelatincapsules it is possible to employ pharmaceutical carriers normally usedfor producing dispersions or suspensions, such as, for example, aqueousgums, celluloses, silicates or oils, which are incorporated into a softgelatin capsule shell. Syrup formulations normally consist of asuspension or solution of the compound or of a salt thereof in a liquidcarrier such as, for example, ethanol, peanut oil, olive oil, glycerolor water, it being possible for flavorings and colorants to be present.

It is possible through topical administration of the inventivecombination of active ingredients to achieve therapeutically effectiveconcentrations even with lower dosages. For this reason, topicalformulations, which include in particular intranasal and inhalationalformulations, are preferred for the purposes of the present invention.

Intranasal preparations may be administered as aqueous or oilysolutions, suspensions or emulsions. For the administration of an activeingredient by inhalation, it can be administered in the form of asuspension, solution or emulsion which is present as dry powder or asaerosol, it being possible to use all customary propellants.

In a preferred embodiment of the invention, the phosphodiesterase-4inhibitor composition is in the form of a nasal spray or of a meteredaerosol or of a metered dry powder for inhalation. The glucocorticoidcomposition is preferably likewise a topical preparation, and for thesoft steroid loteprednol a formulation in the form of nasal spray,metered aerosol or metered dry powder for inhalation is again preferred.

The soft steroid loteprednol etabonate employed according to theinvention is preferably formulated as suspension in water, with furtheringredients such as preservatives, stabilizers, tonicity agents,thickeners, suspension stabilizers, excipients to adjust the pH, buffersystems and wetting agents. For further details of suitable excipients,reference is made for example to DE 19 947 234.

The pharmaceutical preparations of the invention may, besides theglucocorticoid and at least one phosphodiesterase-4 inhibitor activeingredients, comprise further ingredients such as customarypreservatives, stabilizers, thickeners, flavorings, etc.

EXEMPLARY EMBODIMENT

Nasal spray suspension with loteprednol etabonate (1%) Loteprednoletabonate 1.000 g Avicel RC 591 1.100 g Polysorbate 80 0.100 g Sorbitolsolution 70% 6.000 g Sodium edetate 0.050 g Benzalkonium chloride 0.020g Purified water ad 100 mlProduction

Introduce 45 kg of purified water into a suitable agitating containerwith homogenizing device, and homogenize Avicel RC 591 therein at highspeed. Then dissolve the substances polysorbate 80, sorbitol solution,sodium edetate and benzalkonium chloride together while agitating.

Subsequently homogenize the active ingredient loteprednol etabonate athigh speed until a uniform suspension is produced. Then make up thefinal volume with purified water and homogenize further. Subsequentlyevacuate the suspension in order to remove the air bubbles which havebeen produced. The resulting suspension is subsequently dispensed intobottles which are then provided with a suitable nasal spray pump.

In an advantageous embodiment, the active components of this combinationare in the form of a fixed combination, thus simplifying use for thepatient. Administration of the active ingredients can in this case takeplace simultaneously, sequentially or separately in free or fixedcombination. They can be administered both in a single-dose form and astwo separate formulations, which may be identical or different. Deliverycan take place at the same time, simultaneously, or at separate times,by which is meant both short and long intervals such as, for example,administration of loteprednol in the evening and administration of thephosphodiesterase-4 inhibitor in the morning, or vice versa.

The active ingredients can be administered from once to six times a day.The active ingredients are preferably administered once to twice a day,particularly preferably twice a day. The dose of one or morephosphodiesterase-4 inhibitors is approximately from 0.1 to 20 mg perday per adult, preferably between 0.2 and 5 mg. The dose of theglucocorticoid can be in the region of the approved dosage, i.e. in therange from 0.1 to 1.6 mg per day, preferably between 0.2 and 0.8 mg perday. The actual dose depends on the general condition of the patients(age, weight, etc.) and the severity of the disease.

1. A composition comprising a glucocorticoid and at least onephosphodiesterase-4 inhibitor in fixed or free combination.
 2. Thecomposition as claimed in claim 1, characterized in that thephosphodiesterase-4 inhibitor is rolipram, piclamilast, roflumilast,cilomilast, the hydroxyindole derivativeN-(3,5-dichloropyridin-4-yl)-2-[1-(4-fluorobenzyl)-5-hydroxyindol-3-yl]-2-oxoacetamide(DFHO) or their pharmaceutically acceptable salts or mixtures thereof.3. The composition as claimed in 1, characterized in that theglucocorticoid is a soft steroid.
 4. The composition as claimed in claim1, characterized in that the glucocorticoid is beclomethasone,budesonide, ciclesonide, fluticasone, mometasone or loteprednol or apharmaceutically acceptable ester thereof.
 5. The composition as claimedin claim 3, characterized in that the glucocorticoid is loteprednoletabonate.
 6. A medicament for the treatment of respiratory diseases,allergic diseases, asthma and/or chronic obstructive pulmonary diseases,comprising as active ingredient a glucocorticoid and at least onephosphodiesterase-4 inhibitor in fixed or free combination, whereappropriate together with customary excipients or carriers.
 7. Themedicament as claimed in claim 6, characterized in that it can beadministered orally.
 8. The medicament as claimed in claim 6,characterized in that it can be administered topically.
 9. Themedicament as claimed in claim 8, characterized in that it can beadministered simultaneously, sequentially or separately from oneanother, intranasally or by inhalation.
 10. The medicament as claimed inclaim 8, characterized in that it is an inhalable liquid or solidpreparation.
 11. The medicament as claimed in claim 6, characterized inthat one active ingredient is administered orally and at least oneactive ingredient is administered topically.
 12. The medicament asclaimed in claim 6, characterized in that the phosphodiesterase-4inhibitor(s) can be administered orally.
 13. A process for producing amedicament for the treatment and prophylaxis of respiratory diseases,allergic diseases, asthma and/or chronic obstructive pulmonary diseases,comprising as active ingredient a glucocorticoid and at least onephosphodiesterase-4 inhibitor, characterized in that the glucocorticoidand the phosphodiesterase-4 inhibitor(s) are mixed singly or together,where appropriate together with customary excipients and carriers, andthe mixture obtained in this way is converted into suitable dosageforms.
 14. The use of the fixed or free combination of a glucocorticoidand at least one phosphodiesterase-4 inhibitor for producing amedicament for the treatment and prophylaxis of respiratory diseases,allergic diseases, asthma and/or chronic obstructive pulmonary diseases.15. The use as claimed in claim 14, characterized in that theglucocorticoid is loteprednol etabonate and the phosphodiesterase-4inhibitor is the hydroxindole derivativeN-(3,5-dichloropyridin-4-yl)-2-[1-(4-fluorobenzyl)-5-hydroxyindol-3-yl]-2-oxoacetamide(DFHO).